Transactivation of hsp70-1/2 in geldanamycin-treated human non-small cell lung cancer H460 cells: Involvement of intracellular calcium and protein kinase C

✍ Scribed by Chih-Wen Shu; Nai-Lin Cheng; Whei-meih Chang; Tzu-Ling Tseng; Yiu-Kay Lai

• Publisher: John Wiley and Sons
• Year: 2005
• Tongue: English
• Weight: 330 KB
• Volume: 94
• Category: Article
• ISSN: 0730-2312
• DOI: 10.1002/jcb.20348

✦ Synopsis

Abstract

Geldanamycin is an antitumor drug that binds HSP90 and induces a wide range of heat shock proteins, including HSP70s. In this study we report that the induction of HSP70s is dose‐dependent in geldanamycin‐treated human non‐small cell lung cancer H460 cells. Analysis of the induction of HSP70s specific isoform using LC‐ESI‐MS/MS analysis and Northern blotting showed that HSP70‐1/2 are the major inducible forms under geldanamycin treatment. Transactivation of hsp70‐1/2 was determined by electrophoretic mobility‐shift assay using heat shock element (HSE) as a probe. The signaling pathway mediators involved in hsp70‐1/2 transactivation were screened by the kinase inhibitor scanning technique. Pretreatment with serine/threonine protein kinase inhibitors H7 or H8 blocked geldanamycin‐induced HSP70‐1/2, whereas protein kinase A inhibitor HA1004, protein kinase G inhibitor KT5823, and myosin light chain kinase inhibitor ML‐7 had no effect. Furthermore, the protein kinase C (PKC)‐specific inhibitor Ro‐31‐8425 and the Ca^2+^‐dependent PKC inhibitor Gö‐6976 diminished geldanamycin‐induced HSP70‐1/2, suggesting an involvement of the PKC in the process. In addition, geldanamycin treatment causes a transient increase of intracellular Ca^2+^. Chelating intracellular Ca^2+^ with BAPTA‐AM or depletion of intracellular Ca^2+^ store with A23187 or thapsigargin significantly decreased geldanamycin‐transactivated HSP70‐1/2 expression. Taken together, our results demonstrate that geldanamycin‐induced specific HSP70‐1/2 isoforms expression in H460 cells through signaling pathway mediated by Ca^2+^ and PKC. © 2005 Wiley‐Liss, Inc.